Genetic studies have demonstrated that loss of the transcriptional coactivator p/CIP leads to resistance to obesity and diabetes, especially in extreme mouse obesity models. The goal of this research is to find small molecules that decrease p/CIP protein stability, thus providing new understanding and novel approaches to cure obesity and diabetes. In an in vitro white fat differentiation system, p/CIP is important for fat cell formation. A fluorescence-based primary assay employing this in vitro cellular model has been developed to screen for small molecules that destabilize a p/CIP-EGFP fusion protein. Individual compounds will be incubated with the stably transfected cell line containing this GFP (green fluorescence) fusion protein. Small molecules will be identified that only diminish the GFP signal from the p/CIP-EGFP fusion protein cells, but not from a control cell line with GFP protein alone. These candidate compounds therefore increase the fusion protein turnover, and will be examined further for their effects on the endogenous p/CIP protein levels in secondary assays. If the small molecule hits destabilize the endogenous p/CIP protein, they can be used as chemical probes to decipher kinase, as well as proteasome degradation pathways that are critical for controlling p/CIP protein turnover and therefore controlling obesity and diabetes. These compounds will be chemically optimized and tested in readily available mouse obesity models for their effects in vivo. Potential therapeutic drugs may come out of these studies to help alleviate extreme obesity in human patients. PUBLIC HEALTH RELEVANCE: This research will provide crucial new information about how to control obesity and diabetes, and will provide potential cure for the extremely obese human population. [unreadable] [unreadable] [unreadable] [unreadable]